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David M Engman

Publications and source records attributed to David M Engman.

18 recordsLinked to original sources
PubMed2006

The major surface-metalloprotease of the parasitic protozoan, Leishmania, protects against antimicrobial peptide-induced apoptotic killing.

Human infection by the vector-borne protozoan Leishmania is responsible for substantial worldwide morbidity and mortality. The surface-metalloprotease (leishmanolysin) of Leishmania is a virulence factor which contributes to a variety of functions including evasion of complement-mediated parasite-killing and host intramacrophage survival. We tested the hypothesis that leishmanolysin serves to protect parasites from the cytolytic effects of various antimicrobial peptides (AMPs) which are important components of the innate immune system. We found that members of the alpha- and theta-defensins, magainins and cathelicidins had substantially higher leishmanicidal activity against leishmanolysin-knock out mutants of L. major. Using the magainin analogue, pexiganan, as a model peptide we show that AMP evasion is due to rapid and extensive peptide degradation by wild-type parasites. Pexiganan-treatment of knock out mutants induced disruption of surface-membrane permeability and expression of features of apoptosis including smaller cell size, loss of mitochondrial membrane potential, exposure of surface phosphatidyl serine as well as induction of caspase 3/7 activity. These results demonstrate leishmanolysin as a virulence factor preventing AMP-mediated apoptotic killing. This study serves as a platform for the dissection of the AMP-mediated death pathways of Leishmania and demonstrates the potential that AMP evasion plays during host infection by this parasite.

Animals↗
PubMed2006

Induction of cardiac autoimmunity in Chagas heart disease: a case for molecular mimicry.

Up to 18 million of individuals are infected by the protozoan parasite Trypanosoma cruzi in Latin America, one third of whom will develop chronic Chagas disease cardiomyopathy (CCC) up to 30 years after infection. Cardiomyocyte destruction is associated with a T cell-rich inflammatory infiltrate and fibrosis. The presence of such lesions in the relative scarcity of parasites in the heart, suggested that CCC might be due, in part, to a postinfectious autoimmune process. Over the last two decades, a significant amount of reports of autoimmune and molecular mimicry phenomena have been described in CCC. The authors will review the evidence in support of an autoimmune basis for CCC pathogenesis in humans and experimental animals, with a special emphasis on molecular mimicry as a fundamental mechanism of autoimmunity.

Animals↗
PubMed2005

A flagellum-specific calcium sensor.

The flagellar calcium-binding protein (FCaBP) of the flagellated protozoan Trypanosoma cruzi associates with the flagellar membrane via its N-terminal myristate and palmitate moieties in a calcium-modulated, conformation-dependent manner. This mechanism of localization is similar to that described for neuronal calcium sensors, which undergo calcium-dependent changes in conformation, which modulate the availability of the acyl groups for membrane interaction and partner association. To test whether FCaBP undergoes a calcium-dependent conformational change and to explore the role of such a change in flagellar targeting, we first introduced point mutations into each of the two EF-hand calcium-binding sites of FCaBP to define their affinities. Analysis of recombinant EF-3 mutant (E151Q), EF-4 mutant (E188Q), and double mutant proteins showed EF-3 to be the high affinity site (Kd approximately 9 microM) and EF-4 the low affinity site (Kd approximately 120 microM). These assignments also correlated with partial (E188Q), nearly complete (E151Q), and complete (E151Q,E188Q) disruption of calcium-induced conformational changes determined by NMR spectrometry. We next expressed the FCaBP E151Q mutant and the double mutant in T. cruzi epimastigotes. These transproteins localized to the flagellum, suggesting the existence of a calcium-dependent interaction of FCaBP that is independent of its intrinsic calcium binding capacity. Several proteins were identified by FCaBP affinity chromatography that interact with FCaBP in a calcium-dependent manner, but with differential dependence on calcium-binding by FCaBP. These findings may have broader implications for the calcium acyl switch mechanism of protein regulation.

Animals↗
PubMed2005

Responsive microtubule dynamics promote cell invasion by Trypanosoma cruzi.

The American trypanosome, Trypanosoma cruzi, can invade non-phagocytic cell types by a G-protein-mediated, calcium-dependent mechanism, in which the cell's natural puncture repair mechanism is usurped in order to recruit lysosomes to the parasite/host cell junction or 'parasite synapse.' The fusion of lysosomes necessary for construction of the nascent parasitophorous vacuole is achieved by directed trafficking along microtubules. We demonstrate altered host cell microtubule dynamics during the initial stages of the entry process involving de novo microtubule polymerization from the cytoplasmic face of the parasite synapse which appears to serve as a secondary microtubule organizing centre. The net result of these dynamic changes to the host cell's microtubule cytoskeleton is the development of the necessary infrastructure for transport of lysosomes to the parasite synapse.

Animals↗
PubMed2005

Effects of cannabinoid treatment on Chagas disease pathogenesis: balancing inhibition of parasite invasion and immunosuppression.

Trypanosoma cruzi invades heart cells via a calcium-dependent, G protein-mediated mechanism, leading to severe cardiac inflammation considered by some to be autoimmune in nature. Cannabinoids inhibit calcium flux and G protein signalling; as potent immunosuppressive agents, they are effective in the treatment of autoimmune disease but contraindicated for the treatment of infections. We compared the action of the synthetic cannabinoid R(+)WIN55,212 and its inactive isomer S(-)WIN55,212 on cardiac myoblast invasion: R(+)WIN55,212 inhibited invasion by over 85%. We then tested for efficacy in modulating pathogenesis in mice by assaying parasite burden in heart and blood, cellular and humoral immunity to parasite and self antigens, and mortality. R(+)WIN55,212 significantly reduced cardiac inflammation but led to considerably increased parasitaemia. Cardiac parasitosis and mortality were not significantly different in treatment and control groups. We conclude that cannabinoids can block cardiac cell puncture repair mechanisms, thereby inhibiting trypanosome invasion as predicted by the mode of drug action, but, also inhibit immune cell effector functions, offsetting the benefit of inhibition parasite cell invasion. Refined use of cannabinoids may prove therapeutic in the future, but our results raise concern about the effect of cannabis use on those chronically infected by T. cruzi and on heart cell homeostasis generally.

Animals↗
PubMed2004

A cardiac myosin-specific autoimmune response is induced by immunization with Trypanosoma cruzi proteins.

Trypanosoma cruzi is the protozoan parasite that causes Chagas' heart disease, a potentially fatal cardiomyopathy prevalent in Central and South America. Infection with T. cruzi induces cardiac myosin autoimmunity in susceptible humans and mice, and this autoimmunity has been suggested to contribute to cardiac inflammation. To address how T. cruzi induces cardiac myosin autoimmunity, we investigated whether immunity to T. cruzi antigens could induce cardiac myosin-specific autoimmunity in the absence of live parasites. We immunized A/J mice with a T. cruzi Brazil-derived protein extract emulsified in complete Freund's adjuvant and found that these mice developed cardiac myosin-specific delayed-type hypersensitivity (DTH) and autoantibodies in the absence of detectable cardiac damage. The induction of autoimmunity was specific since immunization with extracts of the related protozoan parasite Leishmania amazonensis did not induce myosin autoimmunity. The immunogenetic makeup of the host was important for this response, since C57BL/6 mice did not develop cardiac myosin DTH upon immunization with T. cruzi extract. Perhaps more interesting, mice immunized with cardiac myosin developed T. cruzi-specific DTH and antibodies. This DTH was also antigen specific, since immunization with skeletal myosin and myoglobin did not induce T. cruzi-specific immunity. These results suggest that immunization with cardiac myosin or T. cruzi antigen can induce specific, bidirectionally cross-reactive immune responses in the absence of detectable cardiac damage.

Animals↗
PubMed2003

Myosin autoimmunity is not essential for cardiac inflammation in acute Chagas' disease.

Infection with the protozoan parasite Trypanosoma cruzi leads to acute myocarditis that is accompanied by autoimmunity to cardiac myosin in susceptible strains of mice. It has been difficult to determine the contribution of autoimmunity to tissue inflammation, because other inflammatory mechanisms, such as parasite-mediated myocytolysis and parasite-specific immunity, are coincident during active infection. To begin to investigate the contribution of myosin autoimmunity to myocarditis, we selectively inhibited myosin autoimmunity by restoring myosin tolerance via injection of myosin-coupled splenocytes. This tolerization regimen suppressed the strong myosin-specific delayed-type hypersensitivity (DTH) that normally develops in infected mice, although it did not affect myosin-specific Ab production. Suppression of myosin autoimmunity had no effect on myocarditis or cardiac parasitosis. In contrast, myosin tolerization completely abrogated myocarditis in mice immunized with purified myosin, which normally causes severe autoimmune myocarditis. In this case, myosin-specific DTH and Ab production were significantly reduced. We also examined the contribution of T. cruzi-specific immunity to inflammation by injection of T. cruzi-coupled splenocytes before infection. This treatment reduced T. cruzi DTH, although there was no effect on parasite-specific Ab production. Interestingly, cardiac inflammation was decreased, cardiac parasitosis was significantly increased, and mortality occurred earlier in the parasite-tolerized animals. These results indicate that myosin-specific autoimmunity, while a potentially important inflammatory mechanism in acute and chronic T. cruzi infection, is not essential for inflammation in acute disease. They also confirm previous studies showing that parasite-specific cell-mediated immunity is important for myocarditis and survival of T. cruzi infection.

Acute Disease↗
PubMed2003

Captopril prevents experimental autoimmune myocarditis.

Captopril, an angiotensin-converting enzyme inhibitor, is widely used in the treatment of a variety of cardiomyopathies, but its effect on autoimmune myocarditis has not been addressed experimentally. We investigated the effect of captopril on myosin-induced experimental autoimmune myocarditis. A/J mice, immunized with syngeneic cardiac myosin, were given 75 mg/L of captopril in their drinking water. Captopril dramatically reduced the incidence and severity of myocarditis, which was accompanied by a reduction in heart weight to body weight ratio and heart weight. Captopril specifically interfered with cell-mediated immunity as myosin delayed-type hypersensitivity (DTH) was reduced, while anti-myosin Ab production was not affected. Captopril-treated, OVA-immunized mice also exhibited a decrease in OVA DTH. In myosin-immunized, untreated mice, injection of captopril directly into the test site also suppressed myosin DTH. Interestingly, captopril did not directly affect Ag-specific T cell responsiveness because neither in vivo nor in vitro captopril treatment affected the proliferation, IFN-gamma secretion, or IL-2 secretion by Ag-stimulated cultured splenocytes. These results indicate that captopril ameliorates experimental autoimmune myocarditis and may act, at least in part, by interfering with the recruitment of cells to sites of inflammation and the local inflammatory environment.

Administration, Oral↗
PubMed2003

Killing of African trypanosomes by antimicrobial peptides.

Antimicrobial peptides are components of the innate immune systems of a wide variety of eukaryotic organisms and are being developed as antibiotics in the fight against bacterial and fungal infections. We explored the potential activities of antimicrobial peptides against the African trypanosome Trypanosoma brucei, a vector-borne protozoan parasite that is responsible for significant morbidity and mortality in both humans and animals. Three classes of mammalian antimicrobial peptides were tested: alpha-defensins, beta-defensins, and cathelicidins. Although members of all 3 classes of antimicrobial peptides showed activity, those derived from the cathelicidin class were most effective, killing both insect and bloodstream forms of the parasite. The mechanism of action of the cathelicidins against T. brucei involves disruption of surface membrane integrity. Administration of cathelicidin antimicrobial peptides to mice with late-stage T. brucei infection acutely decreased parasitemia and prolonged survival. These results highlight the potential use of antimicrobial peptides for the treatment of African trypanosomiasis.

Animals↗
PubMed2003

The significance of autoimmunity in the pathogenesis of Chagas heart disease.

Chagas heart disease develops in approximately one-third of individuals infected with the protozoan parasite Trypanosoma cruzi. Among the many possible mechanisms responsible for this illness, an autoimmune mechanism has received much experimental support during the past several decades. Initial observations of the absence or near absence of parasites from the inflamed tissues suggested the autoimmunity hypothesis, and the finding of measurable autoimmune responses in humans and experimental animals has bolstered this idea. The rigorous testing of the hypothesis has been difficult, primarily because other mechanisms are likely to play a role during active infection, particularly immunity to parasite antigens that may persist in the infected animal. While the role autoimmunity plays in disease pathogenesis is not known, it is clear that autoimmune responses are induced during infection of some humans and animals. A number of mechanisms may explain the induction of autoimmunity during T. cruzi infection, including parasite-induced polyclonal lymphocyte activation, molecular mimicry, bystander activation, and presentation of cryptic self epitopes. The genetic makeup of both the parasite and host are also critical to the outcome of infection. The autoimmune hypothesis deserves further exploration, while public health interventions should focus on control of the insects that transmit the parasite, development of parasiticidal drugs and vaccines, and testing of blood products, which have become an important threat of new infections.

Animals↗
PubMed2003

Captopril ameliorates myocarditis in acute experimental Chagas disease.

BACKGROUND: Captopril, an angiotensin-converting enzyme inhibitor, is commonly prescribed to patients with Chagas heart disease (CHD). There are few human studies and no animal studies on the effects of captopril in CHD. We investigated the effects of captopril on myocarditis and the host immune response to Trypanosoma cruzi in an experimental model of acute CHD. METHODS AND RESULTS: A/J mice infected with Brazil strain of T cruzi developed acute myocarditis by day 21 after infection, consisting of severe focal inflammation, necrosis, fibrosis, and T cruzi pseudocysts. Administration of captopril (5 mg/L in the water) significantly reduced necrosis and fibrosis in infected mice. Increasing the captopril dose also led to a decrease in inflammation. Captopril did not affect overall mortality but did delay death while having no effect on parasitemia or cardiac parasite load. Treatment did not affect humoral immunity against T cruzi or cardiac myosin (autoimmunity) but did decrease delayed-type hypersensitivity responses against both antigens. Interestingly, increasing the dose of captopril induced mortality in infected mice in a dose-dependent manner. Mortality was apparently not due to T cruzi because neither parasitemia nor cardiac parasitosis was affected. The combination of captopril and infection may have impaired renal function because these mice had increased water consumption, decreased body mass, and increased serum BUN/creatinine ratio. CONCLUSIONS: Captopril ameliorates the myocarditis associated with acute T cruzi infection.

Acute Disease↗
PubMed2003

A novel reiterated family of transcribed oligo(A)-terminated, interspersed DNA elements in the genome of Trypanosoma cruzi.

We report the molecular characterization of a novel reiterated family of transcribed oligo(A)-terminated, interspersed DNA elements in the genome of Trypanosoma cruzi. Steady-state level of transcripts of this sequence family appeared to be developmentally regulated, since only in the replicative forms the parasite showed expression of related sequences with a major band around 3 kb. The presence of frame shifts or premature stop codons predicts that transcripts are not translated. The sequence family also contains truncated forms of retrotransposons elements that may become potential hot spots for retroelement insertion. Sequences homologous to this family are interspersed at many chromosomes including the subtelomeric regions.

Animals↗
PubMed2003

Migration through the extracellular matrix by the parasitic protozoan Leishmania is enhanced by surface metalloprotease gp63.

Leishmania species engineered to express high levels of the surface metalloprotease gp63 have enhanced capacity of migration through extracellular matrix in vitro. This correlates with gp63 degradation of extracellular matrix components, such as collagen type IV and fibronectin, and suggests an important role for gp63 in the pathogenesis of leishmaniasis.

Animals↗
PubMed2003

Angiotensin converting enzyme inhibitors and angiotensin II receptor antagonists in experimental myocarditis.

Myocarditis is a disease whose pathogenesis is not completely understood and whose prevalence is likely underestimated. Individuals afflicted with this condition may be treated with agents that relieve symptoms arising from inflammation and concurrent cellular damage. One class of drugs commonly used in the treatment of myocarditis includes the angiotensin converting enzyme inhibitors, such as captopril, enalapril and lisinopril, and the angiotensin Pi receptor antagonists, such as L-158,809 and losartan. The effects of these drugs on cardiomyopathy have been studied using a variety of animal models of heart failure and hypertension. However, less research has been done in the area of animal models of frank myocarditis. Here we review the use of angiotensin converting enzyme inhibitors and angiotensin Pi receptor antagonists in animal models of myocarditis. We extend the implications of that published work by correlation with results from studies of other disease models and in vitro experiments that highlight the immunomodulatory potential of these compounds. The literature strongly suggests that aggressive therapy employing angiotensin converting enzyme inhibition and/or blockade of angiotensin Pi receptors is beneficial. Treatment is useful not only for reducing complications associated with myocarditis, but also for downregulating the potential autoimmune component of disease without increasing the levels of the infectious agent that may initiate the myocarditis.

Angiotensin Receptor Antagonists↗
PubMed2002

Extracellular release of the glycosylphosphatidylinositol (GPI)-linked Leishmania surface metalloprotease, gp63, is independent of GPI phospholipolysis: implications for parasite virulence.

The major zinc metalloprotease of Leishmania (gp63), an important determinant of parasite virulence, is attached to the parasite surface via a glycosylphosphatidylinositol anchor. Here we report the spontaneous release of proteolytically active gp63 from a number of Leishmania isolates, causing cutaneous and visceral disease. To investigate the mechanism(s) of gp63 release, we transfected a gp63-deficient variant of Leishmania amazonensis with constructs expressing gp63 and various mutants thereof. Surprisingly, approximately half of wild type gp63 was found in the culture supernatant 12 h post-synthesis. Biochemical analysis of the extracellular gp63 revealed two forms of the protein, one that is released from the cell surface, and another, that apparently is directly secreted. Release of cell surface gp63 was significantly reduced when the proteolytic activity of the protein was inactivated by site-specific mutagenesis or inhibited by zinc chelation, suggesting that release involves autoproteolysis. The extracellular gp63 does not contain a glycosylphosphatidylinositol moiety or ethanolamine, indicating that phospholipolysis is not involved in the release process. Release of gp63 is also independent of glycosylation. The finding of proteolytically active, extracellular gp63 produced by multiple Leishmania isolates suggests a potential role of the extracellular enzyme in substrate degradation relevant to their survival in both the mammalian host and the insect vector.

Animals↗
PubMed2002

Pathogenesis of Chagas heart disease: role of autoimmunity.

Chagas heart disease is caused by infection with the protozoan parasite Trypanosoma cruzi. The apparent absence of parasites from the hearts of most individuals who succumb to this illness has led some to propose an autoimmune basis for disease pathogenesis. This hypothesis has been extremely difficult to test, because other mechanisms of tissue inflammation may coexist in the setting of active infection. Here we review the proposed mechanisms of Chagas disease pathogenesis and present new evidence in support of an autoimmune contribution to cardiac inflammation in the context of these other mechanisms. While we do not yet have a definitive answer to the autoimmunity question, we hope that our views will provide those engaged in the debate fresh perspective on this challenging issue.

Animals↗